RNA sequencing analysis to identify gene targets of Kaposi's sarcoma herpesvirus (KSHV) ORF24

ORF24 is the core component of a virally encoded transcription complex in KSHV that is active during later stages of infection. ORF24 makes interactions with both promoter DNA and host RNA polymerase II. The aim of this study was to identify the targets of the virally encoded transcription complex by sequencing total RNA from cells that express a mutant ORF24 that does not interact with RNA polymerase II and compare it to RNA from a mutant rescue cell line (WT). We observed that most viral genes were down regulated in the absence of the ORF24-Pol II interaction and as expected, majority of them were late genes. Overall design: iSLK cells latently infected with the mutant KSHV ORF24 (RAAAG) or the mutant rescue (MR) were reactivated in to the lytic cycle using doxycycline and sodium butyrate. 24 or 48 hours post reactivation, the total RNA was extracted from the cells and subject to library preparation for RNA sequencing.